Type

Journal Article

Authors

Paula Meleady
Martin Clynes
Orla Coleman
Prashant Kaushik
Martin Power
Michael Henry

Subjects

Biochemistry

Topics
post translational modifications signal transduction cell proliferation quantitative analysis cell division chinese hamster ovary cells cell line cell cycle

Differential Phosphoproteomic Analysis of Recombinant Chinese Hamster Ovary Cells Following Temperature Shift. (2017)

Abstract Phosphorylation is one of the most important post-translational modifications, playing a crucial role in regulating many cellular processes, including transcription, cytoskeletal rearrangement, cell proliferation, differentiation, apoptosis, and signal transduction. However, to date, little work has been carried out on the phosphoproteome in CHO cells. In this study we have carried out a large scale differential phosphoproteomic analysis of recombinant CHO cells following a reduction of culture temperature (temperature shift). The reduction of culture temperature during the exponential phase of growth is commonly employed by the biopharmaceutical industry to increase product yield; however, the molecular mechanisms of temperature shift in CHO cells remain poorly understood. We have identified 700 differentially expressed phosphopeptides using quantitative label-free LC-MS/MS phosphoproteomic analysis in conjunction with IMAC and TiO2 phosphopeptide enrichment strategies, following a reduction in temperature from 37 to 31 °C. Functional assessment of the phosphoproteomic data using gene ontology analysis showed a significant enrichment of biological processes related to growth (e.g., cell cycle, cell division), ribosomal biogenesis, and cytoskeleton organization, and molecular functions related to RNA binding, transcription factor activity, and protein serine/threonine kinase activity. Differential phosphorylation of two proteins, ATF2 and NDRG1, was confirmed by Western blotting. This data suggests the importance of including the post-translational layer of regulation, such as phosphorylation, in CHO "omics" studies. This study also has the potential to identify phosphoprotein targets that could be modified using cell line engineering approaches to improve the efficiency of recombinant protein production.
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Full list of authors on original publication

Paula Meleady, Martin Clynes, Orla Coleman, Prashant Kaushik, Martin Power, Michael Henry

Experts in our system

1
Paula Meleady
Dublin City University
Total Publications: 95
 
2
Martin Clynes
Dublin City University
Total Publications: 209
 
3
Orla Coleman
Dublin City University
Total Publications: 6
 
4
Michael Henry
Dublin City University
Total Publications: 78